1)Peripheral B-lymphocyte deficient A/WySnJ mice vaccinated with the inactivated-virus human diploid cell vaccine(HDCV),the benchmark vaccine for rabies vaccine efficacy, produce low-levels of neutralizing antibody and are not protected against rabies virus challenge. In contrast, A/WySnJ mice vaccinated with live vaccinia recombinant vaccines expressing the rabies virus glycoprotein(G)produce high levels of neutralizing antibody,and are protected. A/WySnJ mice also produce high levels of neutralizing antibody after vaccination with either a rabies DNA vaccine, or a live canarypox vaccine expressing the rabies virus G. The protective efficacy of the DNA and canarypox vaccines against rabies virus challenge will be determined 12 months after vaccination (December, 2004).2) The commercial human HDCV and RabAvert inactivated-rabies virus vaccines reconstituted and held at 4 degrees C for more than one year are as effective as freshly reconstituted vaccines in eliciting neutraling antibody and protecting mice (100%) against rabies virus challenge.3) Beagle dogs vaccinated once intradermally (i.d.) in an ear pinna with a rabies DNA vaccine expressing the rabies virus G produce elevated levels of neutralizing antibody that remain in the protective range for 6 months. In contrast, dogs vaccinated with this vaccine in the quadriceps muscle, via gene gun in the ear pinnae, or i.d. in the neck elicit minimal or undetectable levels of neutralizing antibody. Additional Beagle dogs vaccinated once i.d. in either one or both ear pinna with the DNA vaccine continue to maintain protective levels of neutralizing antibody up to 10 months after vacccination. The levels of antibody are similar to control Beagle dogs that received a one-time intramuscular injection of a commercial canine rabies vaccine. A assay comparing the protective efficacies of the commercial and DNA vaccines will be done at CDC, Atlanta, GA one year after vaccination (November, 2004). At that time, the dogs will be challenged with a salivary gland homogenate obtained from a rabid dog naturally infected with a coyote rabies virus variant.4) Persistent infectious rabies virus and/or rabies virus genome is present in mice 20>500 days after virus injection. Virus and/or genome are primarily detected in bone marrow, injected muscle, and the brain/spinal cord. Rabies virus genome persists in tissues of 30% of immunodeficient mice injected with an avirulent rabies virus. The sensitivity for detecting virus-genome in persistently-infected mice increases 100-fold using real-time Taq-Man PCR analysis. 5)Viremia studies using Taq-Man PCR analysis to detect viral genome show that there are 2 phases of viremia in mice experimentally infected intramuscularly with rabies virus. The initial phase occurs immediately and up to 12 hrs after virus injection. After that time, the genome is only detected in the blood of mice that have developed clinical symptoms of disease such as paralysis.6) Phosphorothioate oligonucleotides (PS-ODNs) are potent inhibitors of in vitro rabies virus replication. The antiviral activity increases with the length of the PS-ODN (40mer>20mer>10mer), and is not dependent on a specific oligonucleotide sequence.